Environmental pollutants can disrupt the homeostasis of endogenous metabolites in organisms, leading to metabolic disorders and syndromes. However, it remains highly challenging to efficiently screen for critical biological molecules affected by environmental pollutants. Herein, we found that enzyme could catalyze hydrogen-deuterium (H-D) exchange between a deuterium-labeled environmental pollutant [D38-bis(2-ethylhexyl) phthalate (D38-DEHP)] and several groups of enzyme-regulated metabolites [cardiolipins (CLs), monolysocardiolipins (MLCLs), phospholipids (PLs), and lysophospholipids (LPLs)]. A high-throughput scanning identified the D-labeled endogenous metabolites in a simple enzyme [phospholipase A2 (PLA2)], enzyme mixtures (liver microsomes), and living organisms (zebrafish embryos) exposed to D38-DEHP. Mass fragmentation and structural analyses showed that similar positions were D-labeled in the CLs, MLCLs, PLs, and LPLs, and this labeling was not attributable to natural metabolic transformations of D38-DEHP or incorporation of its D-labeled side chains. Molecular docking and competitive binding analyses revealed that DEHP competed with D-labeled lipids for binding to the active site of PLA2, and this process mediated H-D exchange. Moreover, competitive binding of DEHP against biotransformation enzymes could interfere with catabolic or anabolic lipid metabolism and thereby affect the concentrations of endogenous metabolites. Our findings provide a tool for discovering more molecular targets that complement the known toxic endpoints of metabolic disruptors.
Keywords: chemical−protein interactions; hydrogen-deuterium exchange; metabolic disorder; metabolic enzyme; zebrafish embryo.